Label-Free Imaging of Mesenchymal Stem Cell Spheroid Differentiation with Flexible-Probe SECM and a Microfluidic Device.

Mesenchymal stem cells (MSCs) have emerged as an indispensable source for stem cell research and preclinical studies due to their capacity for in vitro proliferation and their potential to differentiate into mesodermal lineages, particularly into osteoblasts. This capability has propelled their application in the fields of bone regeneration and osteochondral repair. Traditional methodologies for assessing the differentiation status of MSCs necessitate invasive procedures such as cell lysis or fixation.

Fast and quantitative analysis of level 3 details for latent fingerprints.

Level 3 details play essential roles in practical latent fingerprint (LFP) identification. To reliably extract reproducible and identifiable level 3 features, high-resolution images of fingerprints with adequate quality are required. Conventional methods for acquiring level 3 details often involve specific pretreatment, intricate peripheral, leading to time-consuming analysis.

High-Content Label-Free Single-Cell Analysis with a Microfluidic Device Using Programmable Scanning Electrochemical Microscopy.

The cellular heterogeneity and plasticity are often overlooked due to the averaged bulk assay in conventional methods. Optical imaging-based single-cell analysis usually requires specific labeling of target molecules inside or on the surface of the cell membrane, interfering with the physiological homeostasis of the cell. Scanning electrochemical microscopy (SECM), as an alternative approach, enables label-free imaging of single cells, which still confronts the challenge that the long-time scanning process is not feasible for large-scale analysis at the single-cell level.

Short and long sleeping mutants reveal links between sleep and macroautophagy.

Sleep is a conserved and essential behavior, but its mechanistic and functional underpinnings remain poorly defined. Through unbiased genetic screening in , we discovered a novel short-sleep mutant we named . Positional cloning and subsequent complementation, CRISPR/Cas9 knock-out, and RNAi studies identified Argus as a transmembrane protein that acts in adult peptidergic neurons to regulate sleep.

Antennal and Behavioral Responses of to Volatiles from a Non-Crop Host, .

(Diptera: Drosophilidae) infests a variety of commercial fruits, including cherries and other soft-skinned fruits. After the cropping season of most cultivated crop hosts, it heavily infests the fruit of a wild host-plant, in southwest China. Here, we employ gas chromatography-electroantennographic detection (GC-EAD) together with behavioral bioassays and a trapping experiment to identify volatile semiochemicals emitted by that are involved in attraction.

The Ubiquitin-Specific Peptidase USP18 Promotes Lipolysis, Fatty Acid Oxidation, and Lung Cancer Growth.

Ubiquitin specific peptidase 18 (USP18), previously known as UBP43, is the IFN-stimulated gene 15 (ISG15) deconjugase. USP18 removes ISG15 from substrate proteins. This study reports that USP18-null mice (vs.

Advances in fingermark age determination techniques.

Fingermarks have long been recognized as one of the most reliable and valuable evidence for personal identification. In practice, fingerprint analysis primarily concentrates on latent fingerprint visualization. However, fingerprint visualization techniques do not always enable individualization when fingermarks collected in crime scenes are fragmentary, ambiguous, or deformed.

Hemophilia A ameliorated in mice by CRISPR-based in vivo genome editing of human Factor VIII.

Hemophilia A is a monogenic disease with a blood clotting factor VIII (FVIII) deficiency caused by mutation in the factor VIII (F8) gene. Current and emerging treatments such as FVIII protein injection and gene therapies via AAV-delivered F8 transgene in an episome are costly and nonpermanent. Here, we describe a CRISPR/Cas9-based in vivo genome editing method, combined with non-homologous end joining, enabling permanent chromosomal integration of a modified human B domain deleted-F8 (BDD-F8) at the albumin (Alb) locus in liver cells.

Genetic Mechanisms Underlying Sleep.

Sleep is important for cognitive ability, and perturbations of sleep are associated with a myriad of brain disorders. However, how sleep promotes health and function during wake is poorly understood. To address the cellular and molecular mechanisms underlying sleep, we use the fruit fly as a genetic model.

Systematic Analysis of Different Cell Spheroids with a Microfluidic Device Using Scanning Electrochemical Microscopy and Gene Expression Profiling.

The 3D cell spheroid is an emerging tool that allows better recapitulating of in vivo scenarios with multiple factors such as tissue-like morphology and membrane protein expression that intimately coordinates with enzyme activity, thus providing a psychological environment for tumorigenesis study. For analyzing different spheroids, conventional optical imaging may be hampered by the need for fluorescent labeling, which could cause toxicity side effects. As an alternative approach, scanning electrochemical microscopy (SECM) enables label-free imaging.

Candle Soot Coating for Latent Fingermark Enhancement on Various Surfaces.

We demonstrate a facile method termed candle soot coating (CSC) for fast developing latent fingermarks (LFMs) on various kinds of surfaces (glass, ceramic, metal, paper and adhesive tape). The CSC method can be considered as simple, fast, and low-cost as well as providing high contrast for LFM visualization in potential forensic applications.

Drosophila Nipped-B Mutants Model Cornelia de Lange Syndrome in Growth and Behavior.

Individuals with Cornelia de Lange Syndrome (CdLS) display diverse developmental deficits, including slow growth, multiple limb and organ abnormalities, and intellectual disabilities. Severely-affected individuals most often have dominant loss-of-function mutations in the Nipped-B-Like (NIPBL) gene, and milder cases often have missense or in-frame deletion mutations in genes encoding subunits of the cohesin complex. Cohesin mediates sister chromatid cohesion to facilitate accurate chromosome segregation, and NIPBL is required for cohesin to bind to chromosomes.

Identification of Redeye, a new sleep-regulating protein whose expression is modulated by sleep amount.

In this study, we report a new protein involved in the homeostatic regulation of sleep in Drosophila. We conducted a forward genetic screen of chemically mutagenized flies to identify short-sleeping mutants and found one, redeye (rye) that shows a severe reduction of sleep length. Cloning of rye reveals that it encodes a nicotinic acetylcholine receptor α subunit required for Drosophila sleep.

Interactions between the circadian clock and metabolism: there are good times and bad times.

An endogenous circadian (∼24 h) clock regulates rhythmic processes of physiology, metabolism and behavior in most living organisms. While able to free-run under constant conditions, the circadian clock is coupled to day : night cycles to increase its amplitude and align the phase of circadian rhythms to the right time of the day. Disruptions of the circadian clock are correlated with brain dysfunctions, cardiovascular diseases and metabolic disorders.

FRQ-interacting RNA helicase mediates negative and positive feedback in the Neurospora circadian clock.

The Neurospora circadian oscillator comprises FREQUENCY (FRQ) and its transcription activator, the White Collar Complex (WCC). Repression of WCC's transcriptional activity by FRQ via negative feedback is indispensable for clock function. An unbiased genetic screen that targeted mutants with defects in negative feedback regulation yielded a fully viable arrhythmic strain bearing a novel allele of FRQ-interacting RNA helicase (frh), an essential gene that encodes a putative exosome component protein.

A role for casein kinase 2 in the mechanism underlying circadian temperature compensation.

Temperature compensation of circadian clocks is an unsolved problem with relevance to the general phenomenon of biological compensation. We identify casein kinase 2 (CK2) as a key regulator of temperature compensation of the Neurospora clock by determining that two long-standing clock mutants, chrono and period-3, displaying distinctive alterations in compensation encode the beta1 and alpha subunits of CK2, respectively. Reducing the dose of these subunits, particularly beta1, significantly alters temperature compensation without altering the enzyme's Q(10).

A high-density single nucleotide polymorphism map for Neurospora crassa.

We report the discovery and validation of a set of single nucleotide polymorphisms (SNPs) between the reference Neurospora crassa strain Oak Ridge and the Mauriceville strain (FGSC 2555), of sufficient density to allow fine mapping of most loci. Sequencing of Mauriceville cDNAs and alignment to the completed genomic sequence of the Oak Ridge strain identified 19,087 putative SNPs. Of these, a subset was validated by cleaved amplified polymorphic sequence (CAPS), a simple and robust PCR-based assay that reliably distinguishes between SNP alleles.

A developmental cycle masks output from the circadian oscillator under conditions of choline deficiency in Neurospora.

In Neurospora, metabolic oscillators coexist with the circadian transcriptional/translational feedback loop governed by the FRQ (Frequency) and WC (White Collar) proteins. One of these, a choline deficiency oscillator (CDO) observed in chol-1 mutants grown under choline starvation, drives an uncompensated long-period developmental cycle ( approximately 60-120 h). To assess possible contributions of this metabolic oscillator to the circadian system, molecular and physiological rhythms were followed in liquid culture under choline starvation, but these only confirmed that an oscillator with a normal circadian period length can run under choline starvation.

The band mutation in Neurospora crassa is a dominant allele of ras-1 implicating RAS signaling in circadian output.

band, an allele enabling clear visualization of circadianly regulated spore formation (conidial banding), has remained an integral tool in the study of circadian rhythms for 40 years. bd was mapped using single-nucleotide polymorphisms (SNPs), cloned, and determined to be a T79I point mutation in ras-1. Alterations in light-regulated gene expression in the ras-1(bd) mutant suggests that the Neurospora photoreceptor WHITE COLLAR-1 is a target of RAS signaling, and increases in transcription of both wc-1 and fluffy show that regulators of conidiation are elevated in ras-1(bd).

Enabling a community to dissect an organism: overview of the Neurospora functional genomics project.

A consortium of investigators is engaged in a functional genomics project centered on the filamentous fungus Neurospora, with an eye to opening up the functional genomic analysis of all the filamentous fungi. The overall goal of the four interdependent projects in this effort is to accomplish functional genomics, annotation, and expression analyses of Neurospora crassa, a filamentous fungus that is an established model for the assemblage of over 250,000 species of non yeast fungi. Building from the completely sequenced 43-Mb Neurospora genome, Project 1 is pursuing the systematic disruption of genes through targeted gene replacements, phenotypic analysis of mutant strains, and their distribution to the scientific community at large.

Circadian rhythmicity by autocatalysis.

The temperature compensated in vitro oscillation of cyanobacterial KaiC phosphorylation, the first example of a thermodynamically closed system showing circadian rhythmicity, only involves the three Kai proteins (KaiA, KaiB, and KaiC) and ATP. In this paper, we describe a model in which the KaiA- and KaiB-assisted autocatalytic phosphorylation and dephosphorylation of KaiC are the source for circadian rhythmicity. This model, based upon autocatalysis instead of transcription-translation negative feedback, shows temperature-compensated circadian limit-cycle oscillations with KaiC phosphorylation profiles and has period lengths and rate constant values that are consistent with experimental observations.

Oral vaccination of mice against rodent malaria with recombinant Lactococcus lactis expressing MSP-1(19).

Aim: To construct the recombinant Lactococcus lactis as oral delivery vaccination against malaria.

Methods: The C-terminal 19-ku fragments of MSP1 (MSP-1(19)) of Plasmodium yoelii 265-BY was expressed in L. lactis and the recombinant L.

Effect of F209S Mutation of Escherichia coli AroG on Resistance to Phenylalanine Feedback Inhibition.

In Escherichia coli, 80% of the 3-deoxy-D-arabino-heptulosonate 7-phosphate(DAHP) synthase was encoded by aroG gene. The aroG gene was amplified by polymerase chain reaction(PCR) from strain K-12 and a mutant strain resistant to phenylalanine analogues. The PCR products were cloned and subject to DNA sequence analysis.